Hepatitis C virus (HCV) is a leading cause of chronic hepatitis and hepatocellular carcinoma worldwide and infects more than 1% of the world's population. Successful vaccine development is pivotal in controlling this global health problem. A system for efficient assembly of HCV structural proteins into HCV-like particles (VLPs) in insect cells has been developed in our laboratory. These non-infectious HCV-like particles had similar morphologic, serologic and biophysical properties as the putative virions isolated from HCV infected humans. In contrast to recombinant subunit vaccines, the viral proteins of HCV-like particles may be presented in a native, virion-like conformation and may therefore be superior in eliciting a protective humoral and cellular immune response. Using HCV-LP as a capture antigen in an enzyme-linked immunosorbant assay (ELISA), anti-HCV antibodies were specifically detected in serum from individuals with acute and chronic HCV infection. The ability of the HCV-LP to elicit immune response in vivo was studied in Balb / c mice and New Zealand White rabbits. A specific and broadly directed humoral immune response against the HCV structural proteins with appearance of anti-HCV antibodies was induced. Because of the particular nature of the antigen, cytoxic T cell response, which has been shown to be important in controlling viral infection, may also be induced. Experiments are under way to analyze this aspect of immune response in mice. We plan to conduct experiments in chimpanzees to test the effectiveness of the HCV-LP as a vaccine candidate. Studies have also been extended to full-length HCV cDNA. A full-length HCV genome was assembled from overlapping cDNA clones. HCV polyprotein was expressed from the recombinant baculovirus with correct processing to yield all the structural and nonstructural HCV proteins at high levels. The nonstructural proteins formed complexes in the perinuclear region of the insect cells, possibly as a replicative intermediate. Experiments are underway to determine whether viruslike particles are produced in insect cells and whether the full-length HCV construct is able to replicate and generate infectious virions in this system. These work will not only facilitate studies into HCV replication but also may provide a tissue culture-based system for generation of candidate vaccines and screening of potential antiviral agents.